PROJECT SUMMARY Acute respiratory distress syndrome (ARDS) is a common and formidable complication of critical illness, and is associated with up to 75,000 deaths annually in the United States. Alveolar macrophages (AMs) are the most abundant leukocyte in the resting human lung, and due to their presumed functional heterogeneity and phenotypic plasticity, may play a key role in both the induction and resolution of human ARDS. Current knowledge in animals and in human monocyte-derived macrophages (MDMs) in vitro has shown that macrophages have the ability to modify their structure and function in response to their local microenvironment. However, a major question that remains to be answered is what degree do human AMs undergo reprogramming in ARDS, if at all. The identification of subpopulations of AMs that evolve over the course of ARDS could lead to the development of targeted treatment strategies focused on methods to manipulate AM structure and function in this syndrome. The overall study objective is to determine the role that human AMs play in each phase of ARDS. This project accomplishes the overall study objective by pursuing the following two specific aims: 1) Identify deep immuno- phenotypic profiles in AMs taken from human patients with ARDS by utilizing mass cytometry; and 2) Determine whether the ARDS microenvironment influences AM polarization and function by studying normal human AMs that have been incubated with bronchoalveolar lavage fluid taken from patients at various clinical stages of ARDS. Clarifying the role of AMs in human ARDS is essential to further research in this syndrome. To date, there are no specific pharmacologic therapies directed at the pathogenic mechanisms of ARDS. This project will also directly increase the probability of success for the applicant in becoming an independent translational physician scientist focusing on ARDS and human macrophage biology.